Prostate Specific Membrane Antigen Expression in a Syngeneic Breast Cancer Mouse Model.

PURPOSE: Prostate specific membrane antigen (PSMA) has been studied in human breast cancer (BCa) biopsies, however, lack of data on PSMA expression in mouse models impedes development of PSMA-targeted therapies, particularly in improving breast conserving surgery (BCS) margins. This study aimed to validate and characterize the expression of PSMA in murine BCa models, demonstrating that PSMA can be utilized to improve therapies and imaging techniques. METHODS: Murine triple negative breast cancer 4T1 cells, and human cell lines, MDA-MB-231, MDA-MB-468, implanted into the mammary fat pads of BALB/c mice, were imaged by our PSMA targeted theranostic agent, PSMA-1-Pc413, and tumor to background ratios (TBR) were calculated to validate selective uptake. Immunohistochemistry was used to correlate PSMA expression in relation to CD31, an endothelial cell biomarker highlighting neovasculature. PSMA expression was also quantified by Reverse Transcriptase Polymerase Chain Reaction (RT-PCR). RESULTS: Accumulation of PSMA-1-Pc413 was observed in 4T1 primary tumors and associated metastases. Average TBR of 4T1 tumors were calculated to be greater than 1.5-ratio at which tumor tissues can be distinguished from normal structures-at peak accumulation with the signal intensity in 4T1 tumors comparable to that in high PSMA expressing PC3-pip tumors. Extraction of 4T1 tumors and lung metastases followed by RT-PCR analysis and PSMA-CD31 co-staining shows that PSMA is consistently localized on tumor neovasculature with no expression in tumor cells and surrounding normal tissues. CONCLUSION: The selective uptake of PSMA-1-Pc413 in these cancer tissues as well as the characterization and validation of PSMA expression on neovasculature in this syngeneic 4T1 model emphasizes their potential for advancements in targeted therapies and imaging techniques for BCa. PSMA holds great promise as an oncogenic target for BCa and its associated metastases.

Label-free metabolic imaging for sensitive and robust monitoring of anti-CD47 immunotherapy response in triple-negative breast cancer.

BACKGROUND: Immunotherapy is revolutionizing cancer treatment from conventional radiotherapies and chemotherapies to immune checkpoint inhibitors which use patients' immune system to recognize and attack cancer cells. Despite the huge clinical success and vigorous development of immunotherapies, there is a significant unmet need for a robust tool to identify responders to specific immunotherapy. Early and accurate monitoring of immunotherapy response is indispensable for personalized treatment and effective drug development. METHODS: We established a label-free metabolic intravital imaging (LMII) technique to detect two-photon excited autofluorescence signals from two coenzymes, NAD(P)H (reduced nicotinamide adenine dinucleotide (phosphate) hydrogen) and FAD (flavin adenine dinucleotide) as robust imaging markers to monitor metabolic responses to immunotherapy. Murine models of triple-negative breast cancer (TNBC) were established and tested with different therapeutic regimens including anti-cluster of differentiation 47 (CD47) immunotherapy to monitor time-course treatment responses using the developed metabolic imaging technique. RESULTS: We first imaged the mechanisms of the CD47-signal regulatory protein alpha pathway in vivo, which unravels macrophage-mediated antibody-dependent cellular phagocytosis and illustrates the metabolism of TNBC cells and macrophages. We further visualized the autofluorescence of NAD(P)H and FAD and found a significant increase during tumor growth. Following anti-CD47 immunotherapy, the imaging signal was dramatically decreased demonstrating the sensitive monitoring capability of NAD(P)H and FAD imaging for therapeutic response. NAD(P)H and FAD intravital imaging also showed a marked decrease after chemotherapy and radiotherapy. A comparative study with conventional whole-body bioluminescence and fluorescent glucose imaging demonstrated superior sensitivity of metabolic imaging. Flow cytometry validated metabolic imaging results. In vivo immunofluorescent staining revealed the targeting ability of NAD(P)H imaging mainly for tumor cells and a small portion of immune-active cells and that of FAD imaging mainly for immunosuppressive cells such as M2-like tumor-associated macrophages. CONCLUSIONS: Collectively, this study showcases the potential of the LMII technique as a powerful tool to visualize dynamic changes of heterogeneous cell metabolism of cancer cells and immune infiltrates in response to immunotherapy thus providing sensitive and complete monitoring. Leveraged on ability to differentiate cancer cells and immunosuppressive macrophages, the presented imaging approach provides particularly useful imaging biomarkers for emerged innate immune checkpoint inhibitors such as anti-CD47 therapy.

Immunotherapy for Glioblastoma: Current State, Challenges, and Future Perspectives.

Glioblastoma is the most lethal intracranial primary malignancy by no optimal treatment option. Cancer immunotherapy has achieved remarkable survival benefits against various advanced tumors, such as melanoma and non-small-cell lung cancer, thus triggering great interest as a new therapeutic strategy for glioblastoma. Moreover, the central nervous system has been rediscovered recently as a region for active immunosurveillance. There are vibrant investigations for successful glioblastoma immunotherapy despite the fact that initial clinical trial results are somewhat disappointing with unique challenges including T-cell dysfunction in the patients. This review will explore the potential of current immunotherapy modalities for glioblastoma treatment, especially focusing on major immune checkpoint inhibitors and the future strategies with novel targets and combo therapies. Immune-related adverse events and clinical challenges in glioblastoma immunotherapy are also summarized. Glioblastoma provides persistent difficulties for immunotherapy with a complex state of patients' immune dysfunction and a variety of constraints in drug delivery to the central nervous system. However, rational design of combinational regimens and new focuses on myeloid cells and novel targets to circumvent current limitations hold promise to advent truly viable immunotherapy for glioblastoma.

Integrating the drug, disulfiram into the vitamin E-TPGS-modified PEGylated nanostructured lipid carriers to synergize its repurposing for anti-cancer therapy of solid tumors.

The 'repurposed drug,' disulfiram (DSF), is an inexpensive FDA-approved anti-alcoholism drug with multi-target anti-cancer effect. However, the use of DSF in clinical settings remains limited due to its high instability in blood. In the present study, we created nanostructured lipid carriers (NLC) encapsulated DSF modified with d-α-tocopheryl polyethylene glycol 1000 succinate (vitamin E-TPGS). A spherical shape, superior drug encapsulation (80.7%), and decreased crystallinity of DSF were confirmed with results obtained from TEM, XRD, and DSC analysis. Addition of TPGS considerably improved the physicochemical stability profile of NLC-encapsulated DSF under the different conditions tested here. Furthermore, TPGS-DSF-NLCs outperformed unmodified DSF-NLCs and the free DSF solution by having significantly higher cytotoxicity, lower IC50 value (4T1: 263.2 nM and MCF-7: 279.9 nM), and an enhanced cellular uptake in MCF7 and 4T1 cell lines. In vivo anti-tumor analysis in 4T1 murine xenograft model mice revealed a significant (p-value < 0.05) decrease in tumor volume and higher tumor growth inhibition rate (48.24%) with TPGS-DSF-NLC treatment as compared to both the free DSF solution (8.49%) and DSF-NLC formulations (29.2%). Histopathology analysis of tumor tissues further confirmed a noticeably higher anti-tumor activity of TPGS-DSF-NLC through augmented cell necrosis in solid tumors. Hence, the present study established that addition of TPGS can synergize the anti-cancer activity of NLC-encapsulated DSF formulations, and thus, offer a promising anti-cancer delivery system for DSF.

Interplay between cancer cell cycle and metabolism: Challenges, targets and therapeutic opportunities.

A growing interest has emerged in the field of studying the cross-talk between cancer cell cycle and metabolism. In this review, we aimed to present how metabolism and cell cycle are correlated and how cancer cells get energy to drive cell cycle. Cell proliferation and cell death largely depend on the metabolic activity of the cell. Cell cycle proteins, e.g. cyclin D, cyclin dependent kinase (CDK), some pro-apoptotic and anti-apoptotic proteins, and P53 have been shown to be regulated by metabolic crosstalk. Dysregulation of this cross-talk between metabolism and cell cycle leads to degenerative disorder(s) and cancer. It is not fully understood the actual reason of aberration between metabolism and cell cycle, but it is a hallmark of cancer research. Herein, we discussed the role of some regulatory molecules relative of cell cycle and metabolism and highlight how they control the function of each other. We also pointed out, current therapeutic opportunities and some additional crucial therapeutic targets on these fields that could be a breakthrough in cancer research.

Influence of Tumor Microenvironment on the Distribution and Elimination of Nano-formulations.

OBJECTIVE: To give an in-depth overview about the tumor and its surrounding microenvironment influencing distribution and elimination of nanoformulations. Mehtods: This up-to-date review will summarize the microenvironmental components and their influence on the various factors related to nanoformulations and tumor which affect the penetration, distribution, regulation and clearance of nanoformulations from the tumor cells. Results of recent advances in miroenvironmental tuning with nanoformulations will be evaluated mechanistically. In addition, those natures of tumors involving enhanced cancer therapy will be discussed. Finally, strategies of nanoparticulate design and decoration to achieve efficient drug delivery are presented. RESULTS: Development of tumor is facilitated by its surrounding microenvironment and is regulated by different extra and intracellular components. Drug-loaded nanoformulations are mainly administered via oral and parenteral routes which reach tumor cells via different mechanisms. Chemotherapeutics get diffused from circulation into the surrounding microenvironment which latter get internalized into the cellular interstial area by passive diffusion mechanism due to virtue of size, charge, and pegylation effects, or by ligands and receptor mediated, or through enhanced permeability and retention effects through leaky apertures. Due to mildly acidic environment and hypoxic interstial environment, the influx of nanoformulations is hindered. The metabolites of the nanoformulations get diffused out from the tumor cells as a results of high interstial fluid pressure and get cleared either via liver or via renal execration. CONCLUSION: Well-understanding tumoral microenvironments which significantly affect distribution and elimination of nanoformulations is essential for engineering delivery systems with superior anti-tumoral effect.